Live cell imaging and other cell culture monitoring is widely used in bioscience experiments for a better understanding of dynamic cellular behaviors such as migration, division, differentiation, interaction with the environment, and organelle-level events. Cell culture monitoring can also provide the opportunity to detect unanticipated cell death and contamination at an early stage to rescue failing cell culture in a timely manner.
For longitudinal studies, bioscientists have traditionally resorted to building a specialized incubation and imaging chamber onto a microscope, and imaging the cultured cells directly on the microscope stage as discussed in http://www.zeiss.com/microscopy/en_us/products/microscope-components/incubation.html. This approach, however, is not only expensive, but occupies a significant amount of real estate in the lab. Furthermore, field-of-view and image resolution are coupled to the physical properties of the objective lens in the microscope. Thus field-of-view and image resolution must be traded off when using this microscope platform. Another approach to cell culture monitoring is to incorporate imaging systems within incubators. Unfortunately, these systems also employ a standard microscope. Moreover, mechanical scanning is used. As a result, it is difficult to obtain wide field of view and high resolution imaging at the same time in these systems. In addition, these systems are expensive to build and maintain and their throughput is limited by the standard microscope itself. That is, the number of resolvable pixels, characterized by the space-bandwidth product accessible through a standard microscope is typically limited to 10 megapixels. This space-bandwidth product limitation constrains the rate of image acquisition or throughput achieved by the systems.
Some on-chip microscopes have been developed with the purpose of overcoming the space-bandwidth product limit of the standard microscope. These on-chip microscopes have demonstrated successful high resolution and wide field-of-view imaging of the cell cultures from within the incubator. Examples of these on-chip microscopes can be found at G. Zheng, S. A. Lee, Y. Antebi, M. B. Elowitz, and C. Yang, “The ePetri dish, an on-chip cell imaging platform based on subpixel perspective sweeping microscopy (SPSM),” Proc. Natl. Acad. Sci. U.S.A. 108(41), 16889-16894 (2011), J. H. Jung, C. Han, S. A. Lee, J. Kim, and C. Yang, “Microfluidic-integrated laser-controlled microactuators with on-chip microscopy imaging functionality,” Lab Chip 14(19), 3781-3789 (2014), and C. Han, S. Pang, D. V. Bower, P. Yiu, and C. Yang, “Wide field-of-view on-chip Talbot fluorescence microscopy for longitudinal cell culture monitoring from within the incubator,” Anal. Chem. 85(4), 2356-2360 (2013). However, these on-chip microscopes have the inherent limitation that cells need to be grown on top of the image sensor. This limitation is a marked departure from the standard cell culture workflow. If culture cells or biological sample is imaged on top of the sensor chip, the surface is made with silicon-based material (usually silicon nitride), so cell culture environment changes. Even though a special layer may be coated onto the image sensor, the bottom layer of the surface is different from plastic or glass used in a standard cell culture workflow. Furthermore, the imaging sensor surface is an active layer and making heat during operation. So the cell culture environment can be affected by the temperature change related with this heat unless the system is designed for the cooling. Other lensless imaging methods, such as digital in-line holography, can work without this restriction and can provide high imaging space-bandwidth product in the bright-field mode, but the absence of optical focusing elements prevents them from having effective fluorescence imaging capability. Focusing elements are needed for effective fluorescence imaging capability since fluorescence emissions are incoherent and of low intensity, which degrades the resolution of fluorescence images without any focusing elements. Examples of digital in-line holography can be found in W. Bishara, T. W. Su, A. F. Coskun, and A. Ozcan, “Lensfree on-chip microscopy over a wide field-of-view using pixel super-resolution,” Opt. Express 18(11), 11181-11191 (2010), W. Bishara, U. Sikora, O. Mudanyali, T. W. Su, O. Yaglidere, S. Luckhart, and A. Ozcan, “Holographic pixel super-resolution in portable lensless on-chip microscopy using a fiber-optic array,” Lab Chip 11(7), 1276-1279 (2011), and A. Greenbaum, U. Sikora, and A. Ozcan, “Field-portable wide-field microscopy of dense samples using multi-height pixel super-resolution based lensfree imaging,” Lab Chip 12(7), 1242-1245 (2012).